Effects of Tribulus terrestris saponins on proliferation and invasion of A549 cells

Objective:Tribulus terrestris saponin is a traditional Chinese medicine in China.This experiment was designed to investigate the effects of tribulus terrestris saponin on the proliferation and invasion ability of non-small cell lung cancer A549 cells.Methods:A549 cells were divided into normal control and experimental groups(Tribulus terrestris saponin 250μg/mL group,Tribulus terrestris saponin 200μg/mL group,Tribulus terrestris saponin 150μg/mL group,Tribulus terrestris saponin 100μg/mL group,Tribulus terrestris saponin 50μg/mL group).The proliferation viability of the cells in each group was detected by CCK8,the invasion of tumor cells was detected by Transwell model.The mRNA expression of MMP9 and caspase-3 in each group of cells was detected by RT-PCR.Immunofluorescence staining was used to observe the fluorescence intensity of caspase-3 in each group of cells.Results:Compared with the normal control group,tribulus terrestris saponin significantly inhibited the proliferation activity and invasion ability of A549 cells,which was statistically significant(P<0.01).In the invasion assay,compared with the control group,MMP9 expression was significantly reduced and caspase-3 expression was significantly increased in the tribulus terrestris saponin group,and both were concentration-dependent,with statistically significant differences(P<0.01).By cellular immunofluorescence staining experiments,it was found that the fluorescence expression of caspase-3 was enhanced in the experimental group compared with the normal control group,in which the high concentration saponin group was significantly higher than the low concentration group.Conclusion:Tribulus terrestris saponin can inhibit the invasive ability of A549 cells by down-regulating the expression of MMP9,and induce irreversible apoptosis by up-regulating the activation of caspase-3 expression to form caspase-3.

Antibacterial and antifungal activities of different parts of Tribulus terrestris L. growing in Iraq

Antimicrobial activity of organic and aqueous extracts from fruits, leaves and roots of Tribulus terrestris L., an Iraqi medicinal plant used as urinary anti-infective in folk medicine, was examined against I l species of pathogenic and non-pathogenic microorganisms： Staphylococcus aureus, Bacillus subtilis, Bacillus cereus, Corynebacterium diphtheriae, Escherichia coli, Proteus vulgaris, Serratia marcescens, Salmonella typhimurium, Klebsiella pneumoniae, Pseudomonas aeruginosa and Candida albicans using microdilution method in 96 multiwell microtiter plates. All the extracts from the different parts of the plant showed antimicrobial activity against most tested microorganisms. The most active extract against both Gram-negative and Gram-positive bacteria was ethanol extract from the fruits with a minimal inhibitory concentration （MIC） value of 0.15 mg/ml against B. subtilis, B. cereus, P. vulgaris and C. diphtheriae. In addition, the same extract from the same plant part demonstrated the strongest antifungal activity against C. albicans with an MIC value of 0.15 mg/ml.

Tribulus terrestris extracts alleviate muscle damage and promote anaerobic performance of trained male boxers and its mechanisms： Roles of androgen, IGF-1, and IGF binding protein-3

Purpose: To investigate the effects of Tribulus terrestris(TT) extracts on muscle mass, muscle damage, and anaerobic performances of trained male boxers and its mechanisms: roles of plasma androgen, insulin growth factor 1(IGF-1), and IGF-1 binding protein-3(IGFBP-3).Methods: Fifteen male boxers were divided into exercise group(E, n = 7) and exercise plus TT group(E + TT, n = 8). The 2 groups both undertook3-week high-intensity and 3-week high-volume trainings separated by a 4-week rest. TT extracts(1250 mg/day) were orally administered by boxers in E + TT group. TT extract compositions were detected by UHPLC–Q-TOF/MS. Before and at the end of the 2 trainings, muscle mass, anaerobic performance, and blood indicators were explored.Results: Compared with E group, decreases of plasma CK(1591.5 ± 909.6 U/L vs. 2719.9 ± 832.5 U/L) and IGFBP-3(3075.5 ± 1072.5 ng/m L vs. 3950.8 ± 479.3 ng/m L) as well as increases of mean power(MP, 459.4 ± 122.3 W vs. 434.6 ± 69.5 W) and MP/body weight(MP/BW, 7.5 ± 0.9 W/kg vs. 7.1 ± 1.1 W/kg) were detected in E + TT group after a high-intensity training. For high-volume training, reduction of IGFBP-3(2946.4 ± 974.1 ng/m L vs. 3632.7 ± 470.1 ng/m L) and increases of MP(508.7 ± 103.2 W vs. 477.8 ± 49.9 W) and MP/BW(8.2 ± 0.3 W/kg vs.7.5 ± 0.9 W/kg) were detected in E + TT group, compared with E group. Muscle mass, blood levels of testosterone, dihydrotestosterone(DHT),and IGF-1 were not signifiantly changed between the 2 groups.Conclusion: Taking 1250 mg capsules containing TT extracts did not change muscle mass and plasma levels of testosterone, DHT, and IGF-1 but significantly alleviated muscle damage and promoted anaerobic performance of trained male boxers, which may be related to the decrease of plasma IGFBP-3 rather than androgen in plasma.

Terresoxazine,A Novel Compound with Benzoxazine Skeleton from Tribulus terrestris

Terresoxazine, a novel benzoxazine derivative was isolated from the fruits of Tribulus terrestris. Its structure was determined as 7-hydroxy-3, 3a-dihydro-5H-pyrrolo-[1,2-a] [3,1] -benzoxazin-1(2H)-one, on the basis of the spectral techniques and X-ray crystallographic analysis.

An Acidic Polysaccharide from Tribulus terrestris

An aqueous acidic polysaccharide, named rhamnogalacturonan (designated as TTP-D2) was isolated from Tribulus terrestris L by means of DEAE-cellulose chromatography and gel filtration. The molecular mass of TTP-D2 was estimated to be 26 KDa by gel filtration. TTP-D2 is composed of galacturonic acid, rhamnose, arabinose, galactose, fucose, mannose, xylose and glucose in a ratio of 71.4 : 13.5 : 5.6 : 4.9 : 3.1 : 1.9 : 1.9 : 1.0. The main chain structure of TTP-D2 was elucidated as an acidic hetero-polysaccharide with the connection of a-(1-4) galacturonic acid with a-(1-3) rhamnose by GC analysis of partially hydrolyzed products and the determination of 1H, 13C-NMR spectra.

Two New Furostanol Saponins from Tribulus Terrestris L.

Two new furostanol glucosides, named tribufurosides G and H, were isolated from Tribulus terrestris L. The structures of the two new furostanol glucosides were elucidated as 26-O-β-D-glucopyranosyl-（25R）-5α-furost12-one-3β,22α,26-tetraol-3-O-β-D-glucopyranosyl-（ 1→2 ）-β-D-glucopyranosyl-（ 1→4 ）-β-D-galactopyranoside（ 1 ） and 26-O-β-D-glucopyranosyl-（25S）-5 α- 12-one-2 α,3,8,22 α,26-tetraol-3 -O-β-D-glucopyranosyl-（ 1 → 2 ）-β-D-glucopyranosyl-（1→4）-β-D-galactopyranoside（2） by 1 D, 2D NMR techniques, ESI-MS analysis as well as chemical methods.

Two new furostanol saponins from Tribulus terrestris L.

Two new furostanol glycoside,26-O-β-D-glucopyranosyl-(25S)-5α-furost-3β,22α,26-triol-3-O-β-D-rhamnopyranosyl-(1→2)-[β-D-glucopyranosyl-(1→4)]-β-D-galactopyranoside(1) and 26-O-β-D-glucopyranosyl-(25S)-5α-furost-20(22)-en-2α,3β,26-triol-3-O-β-D-glucopyranosyl-(1→2)-O-β-D-glucopyranosyl-(1→4)-β-D-galactopyranoside(2) were isolated fromthe fruits of Tribulus terrestris L.The structures of two new furostanol saponins were elucidated by spectroscopic methods.

Use of a Natural Compound Made of Ecklonia bicyclis Seaweed, Tribulus terrestris and Water-Soluble Chitosan Oligosaccharide, in Male Sexual Asthenia with Mild or Mild-Moderate Erectile Dysfunction and Serum Testosterone Levels at the Lower Limit of Normal

Objectives: to evaluate the effectiveness of a natural compound made of Ecklonia bicyclis Seaweed, Tribulus terrestris and water-soluble chitosan oligosaccharide, in the male sexual asthenia with mild or mild-moderate erectile dysfunction and serum testosterone levels between 280 and 350 ng/dl. Materials and Methods: 84 male patients affected by reduced libido and serum testosterone levels at the lower limit of normal, were recruited. We have separated patients in three different age groups: group A (18 - 45 years), group B (45 - 59 years), group C (>60 years). All subjects answered the International index of erectile function questionnaire (IIEF-5) and underwent determination of serum total testosterone before and after 30 days of treatment. Results: Before treatment, the group A showed mean (± standard deviation) total testosterone 321.9 ± 19.2 ng/dl and mean IIEF-5 18.6 ± 1.97, in the group B it was 318.5 ± 18.1 ng/dl and 16.3 ± 2.66, and finally in the group C it was 305.4 ± 13.1 ng/dl and 14.2 ± 1.95 respectively. After treatment mean total testosterone and mean IIEF-5 were respectively: group A (448 ± 111.46 ng/dl and 21.84 ± 3.41);group B (453.8 ± 105.23 ng/dl and 20.4 ± 3.81);group C (385.8 ± 87.29 ng/dl and 16.7 ± 3.84). Conclusions: The treatment with Ecklonia bicyclis, Tribulus terrestris and water-soluble chitosan oligosaccharide might represent a safe and effective option on the improvement of libido and erectile function in man with testosterone level at the lower limit of normal.

Steroidal saponins with anti-inflammatory activity from Tribulus terrestris L.

Objective:Tribulus terrestris L.(T.terrestris)is a highly valuable traditional Chinese medicine used to treat stroke,inflammation,pulmonary fibrosis,liver cancer,and urolithiasis.To identify the basic substance responsible for the anti-inflammatory effect of TST(total saponins of Tribulus),its chemical composition was systematically studied,and its effect of inhibiting nitric oxide generation and the expression of related inflammatory factors were determined.Methods:To separate chemical constituents from T.terrestris by column chromatography.Spectroscopic methods,including 1D and 2D nuclear magnetic resonance spectroscopy(NMR)and mass spectrometry(MS)techniques,were used to elucidate the isolated compounds.The anti-inflammatory activities of TST and several compounds were evaluated in vitro.Results:Fifteen steroidal saponins,including 9 furostanol steroidal saponins(1,2,3,4,5,6,7,8,and 15)and 6 isospirostanol steroidal saponins(9,10,11,12,13,and 14),were isolated from T.terrestris.TST significantly decreased the expression of tumor necrosis factor-a(TNF-a)and interleukin-6(IL-6)in RAW 264.7 cells stimulated by lipopolysaccharides.Compounds 13 and 15 evidently reduced TNF-a expression.Compounds 6,10,12,13,and 15 markedly reduced IL-6 expression.Conclusions:Compounds 1 was a novel furostanol steroidal saponin,named 26-O-b-D-glucopyranosyl-(25R)-5afurostan-12-carbonyl-20(22)-en-3b,26-diol-3-O-{b-D-xylopyranosyl-(1→2)-[b-D-xylopyranosyl-(1→3)]-b-D-glucopyranosyl-(1→4)-[a-L-rhamnopyranosyl-(1→2)]-b-D-galactopyranoside}.Compounds 2 was isolated from the family Zygophyllaceae for the first time,and 5 and 6 were isolated from the Tribulus genus.TST and compounds 6,10,12,13,and 15 exerts antiinflammatory activity.

Separation and Bio-activities of Spirostanol Saponin from Tribulus terrestris

Gross saponins of Tribulus terrestris（GSTT） have exact effect on cardiovascular and cerebrovascular diseases.But as a mixture,the specific efficient component of GSTT is still unknown.Nine monomers of spirostanol saponins were isolated and idendified as JA―JI（named transitorily） by means of NMR spectrometry.After bio-activity screening on them,we defined that monomers tigogenin 3-O-β-D-xylopyranosyl（1→2）-[β-D-xylopyranosyl（1→3）]-β-D-glucopyranosyl（1→4）-[α-L-rhamnopyranosyl（1→2）]-β-D-galactopyranoside（compound JB） and hecogenin3-O-β-D-glucopyranosyl（1→4）-β-D-galactopyranoside（compound JG） have cytoprotective bio-activity.Compound JB display effective dose in 10-8 and 10-9 mol/L,and JG in 10-6―10-9 mol/L.Survival rate,lactate dehydrogenase（LDH） and apoptosis also show that JB（at dose 10-8,10-9 and 10-10 mol/L） can protect myocardial injury caused by hypoxia/reoxygenation（H/R）.While morphology change also shows JG has cytoprotective bio-activity.

Synergistic Hepatoprotective Effect of Feronia fimonia L,, Citrullus colocynthis L,, and Tribulus terrestris L, against Paracetamol Induced Hepatotoxicity in Swiss Albino Rats

Paracetamol toxicity induced a significant rise in AST （aspartate transaminase）, ALT （alanine transaminase）, ALP （alkhaline phisphatase）, total bilirubin, total cholesterol, Triglycerides. Administration of 150 mg/kg b.w of synergistic ethanolic extracts of Feronia limonia L., Citrullus colocynthis L., and Tribulus terrestris L. effectively reduced these pathological damages caused by paracetamol intoxication. In addition to serum parameters treatment of 150 rng/kg b.w of synergistic ethanolic extract of Feronia limonia L., Citrullus colocynthis L. and Tribulus terrestris L. also promote the body weight in albino rats. Histopathological changes of the liver samples were compared with the normal control. From our results, we may infer that the mode of action of synergistic 90% ethanolic extracts of Feronia limonia L., Citrullus colocynthis L., and Tribulus terrestris L. （150 mg/kg b.w） in affording the in vivo hepatoprotective activity against paracetamol may be due to the cell membrane stabilization, hepatic cell regeneration and activation of antioxidative enzymes such as glutathione reductase, glutathione peroxidase, superoxide dismutase and catalase.

What is the effect of Tribulus terrestris on testicular morphology and sperm production?An experimental study in the rat model

The objective of this study was to investigate whether treatment with Tribulus terrestris(Tt)has any impact on the testicular morphology and function in a rodent model.Twenty male rats were divided into a control group and a group receiving 100 mg kg^(-1) body weight of Tt supplementation.After 4o days of experiment,the animals were submitted to euthanasia;epididymal tail spermatozoa were collected;and spermatozoa concentration,motility,and viability were analyzed.In addition,testicles were collected and processed for histomorphometrical analyses.Data were compared using the Student's t-test and considered significant when P<O.05.Spermatozoa concentration,motility,and viability showed no difference between the groups.Further,testicular weight and volume,seminiferous tubule diameter,tunica propria surface density,seminiferous epithelium surface density,and intertubular compartment surface density were statistically similar between the groups.However,seminiferous epithelium height and tubular lumen surface density were augmented in animals treated with Tt.Treatment with Tt does not cause a major impact on testicular morphology,promoting only subtle modifications.No difference on spermatozoa parameters was observed.

Vascular protective effects of aqueous extracts of Tribulus terrestris on hypertensive endothelial injury

Angiotensin Ⅱ(Ang Ⅱ) is involved in endothelium injury during the development of hypertension. Tribulus terrestris(TT) is used to treat hypertension, arteriosclerosis, and post-stroke syndrome in China. The present study aimed to determine the effects of aqueous TT extracts on endothelial injury in spontaneously hypertensive rats(SHRs) and its protective effects against Ang Ⅱ induced injury in human umbilical vein endothelial cells(HUVECs). SHRs were administered intragastrically with TT(17.2 or 8.6 g·kg^(-1)·d^(-1)) for 6 weeks, using valsartan(13.5 mg·kg^(-1)·d^(-1)) as positive control. Blood pressure, heart rate, endothelial morphology of the thoracic aorta, serum levels of Ang Ⅱ, endothelin^(-1)(ET^(-1)), superoxide dismutase(SOD) and malonaldehyde(MDA) were measured. The endothelial injury of HUVECs was induced by 2 × 10–6 mol·L^(-1) Ang Ⅱ. Cell Apoptosisapoptosis, intracellular reactive oxygen species(ROS) was assessed. Endothelial nitric oxide synthase(eN OS), ET^(-1), SOD, and MDA in the cell culture supernatant and cell migration were assayed. The expression of hypertension-linked genes and proteins were analyzed. TT decreased systolic pressure, diastolic pressure, mean arterial pressure and heart rate, improved endothelial integrity of thoracic aorta, and decreased serum leptin, Ang Ⅱ, ET^(-1), NPY, and Hcy, while increased NO in SHRs. TT suppressed Ang Ⅱ-induced HUVEC proliferation and apoptosis and prolonged the survival, and increased cell migration. TT regulated the ROS, and decreased mR NA expression of Akt1, JAK2, PI3Kα, Erk2, FAK, and NF-κB p65 and protein expression of Erk2, FAK, and NF-κB p65. In conclusion, TT demonstrated anti-hypertensive and endothelial protective effects by regulating Erk2, FAK and NF-κB p65.

Aqueous Extracts of Tribulus terrestris Protects Against Oxidized Low-Density Lipoprotein-Induced Endothelial Dysfunction

Objective： To investigate the role of aqueous extracts of Tribulus terrestris （TT） against oxidized low-density lipoprotein （ox-LDL）-induced human umbilical vein endothelial cells （HUVECs） dysfunction in vitro. Methods： HUVECs were pre-incubated for 60 min with TT （30 and 3 μg/mL respectively） or 105 mol/L valsartan （as positive controls） and then the injured endothelium model was established by applying 100 μg/mL ox-LDL for 24 h. Cell viability of HUVECs was observed by real-time cell electronic sensing assay and apoptosis rate by Annexin V/PI staining. The cell migration assay was performed with a transwell insert system. Cytoskeleton remodeling was observed by immunofluorescence assay. The content of endothelial nitric oxide synthase （eNOS） was measured by enzyme-linked immunosorbent assay. Intracellular reactive oxygen species （ROS） generation was assessed by immunofluorescence and flow cytometer. Key genes associated with the metabolism of ox-LDL were chosen for quantitative real-time polymerase chain reaction to explore the possible mechanism of TT against oxidized LDL-induced endothelial dysfunction. Results： TT suppressed ox-LDL-induced HUVEC proliferation and apoptosis rates significantly （41.1% and 43.5% after treatment for 3 and 38 h, respectively; P〈0.05）. It also prolonged the HUVEC survival time and postponed the cell＇s decaying stage （from the 69th h to over 100 h）. According to the immunofluorescence and transwell insert system assay, TT improved the endothelial cytoskeletal network, and vinculin expression and increased cell migration. Additionally, TT regulated of the synthesis of endothelial nitric oxide synthase and generation of intracellular reactive oxygen species （P〈0.05）. Both 30 and 3μg/mL TT demonstrated similar efficacy to valsartan. TT normalized the increased mRNA expression of PI3Kα and Socs3. It also decreased mRNA expression of Aktl, AMPKα, JAK2, LepR and STAT3 induced by ox-LDL. The most notable changes were JAK2, LepR, PI3Kα, Socs3 and STAT3. Conclusions： TT demonstrated potential lowering lipid benefits, anti-hypertension and endothelial protective effects. It also suggested that the JAK2/STAT3 and/or PI3K/AKT pathway might be a very important pathway which was involved in the pharmacological mechanism of TT as the vascular protective agent.

Protection by the gross saponins of Tribulus terrestris against cerebral ischemic injury in rats involves the NF-κB pathway

The aim of this study was to investigate whether the gross saponins of Tribulus terrestris(GSTT),a traditional Chinese herbal medicine,have neuroprotective effects on rats subjected to middle cerebral artery occlusion(MCAO),through nuclear factor-κB(NF-κB)pathway and inflammatory mediators.Cerebral ischemia was produced by MCAO in either untreated(control)or GSTT-pretreated rats,and the animals were examined for infarct volume,cerebral edema,neuro-behavioral abnormality and pathological changes.Meanwhile,the expression of NF-kB protein in brain tissue was analyzed on Western blots and the serum levels of TNF-α and IL-1 were determined by ELISA.The experimental results demonstrated that,compared with the control MCAO group,GSTT-pretreated MCAO group had significantly reduced infarct volume,brain edema and neuro-behavioral abnormality,and lesser degree of pathologic changes in the brain,as well as had lower levels of serum TNF-α and IL-1β,and higher levels of brain NF-κB(Po0.05).Furthermore,treatment with an NF-κB inhibitor pyrrolidine dithiocarbamate(PDTC)abolished the protective effects of GSTT against MCAO-induced cerebral ischemic injury.These results indicated that GSTT’s ability to protect against cerebral ischemic injury was mediated through the NF-κB signaling pathway,and that GSTT may act through inhibition of the production of inflammatory mediators.

Antiglycation and antioxidant activities of the crude extract and saponin fraction of Tribulus terrestris before and after microcapsule release

Objective:The present study investigated antiglycation and antioxidant activities of crude dry extract and saponin fraction of Tribulus terrestris.It also developed a method of microencapsulation and evaluated antiglycation and antioxidant activities of crude dry extract and saponin fraction before and after microcapsule release.Methods:Antiglycation activity was determined by relative electrophoretic mobility(REM),free amino groups and inhibition of advanced glycation end-product(AGE)formation.Antioxidant activity was determined by 2,2-diphenyl-1-picrylhydrazyl(DPPH),ferric ion-reducing antioxidant power(FRAP),nitric oxide(NO)and thiobarbituric acid reactive species(TBARS)tests.Microcapsules were prepared using maltodextrin as wall material and freeze-drying as encapsulation technique.Morphological characterization of microcapsules was evaluated by scanning electron microscopy,and encapsulation efficiency and microcapsule release were determined by total saponins released.Antiglycation and antioxidant assays were performed using crude dry extract and saponin fraction of T.terrestris before and after release.Results:Saponin fraction showed an increase of 32.8%total saponins.High-performance liquid chromatography-mass spectrometry analysis showed the presence of saponins in the obtained fraction.Antiglycation evaluation by REM demonstrated that samples before and after release presented antiglycation activity similar to bovine serum albumin treated with aminoguanidine.Additionally,samples inhibited AGE formation,highlighting treatment with saponin fraction after release(89.89%).Antioxidant tests demonstrated antioxidant activity of the samples.Crude dry extract before encapsulation presented the highest activities in DPPH(92.00%)and TBARS(32.49%)assays.Saponin fraction before encapsulation in FRAP test(499μmol Trolox equivalent per gram of dry sample)and NO test(15.13μmol nitrite formed per gram of extract)presented the highest activities.Conclusion:This study presented antiglycation activity of crude dry extract and saponin fraction of T.terrestris,besides it demonstrated promising antioxidant properties.It also showed that the encapsulation method was efficient and maintained biological activity of bioactive compounds after microcapsule release.These results provide information for further studies on antidiabetic and antiaging potential,and data for new herbal medicine and food supplement formulations containing microcapsules with crude extract and/or saponin fraction of T.terrestris.

An overview of benign prostatic hyperplasia and its appreciation in Greco-Arab (Unani) system of medicine

Objective:Conventional treatments for benign prostatic hyperplasia(BPH)like 5alpha-reductase inhibitors and invasive surgery are associated with some obvious side effects.Conversely,evidence,though limited,has shown that alternative medicines are safer and have potential to improve the lower urinary tract symptoms(LUTS)and quality of life in addition to improving sexual dysfunction in patients with BPH.The current article aimed to include an overview of BPH,different ways of its management,and particularly its appreciation in Greco-Arab(Unani)system of medicine,one of the alternative medicinal systems.Methods:PubMed,Scopus,ScienceDirect,Web of Sciences,Google Scholar databases and classical texts of Greco-Arab medicine were searched for data collection.Results:In Unani system of medicine,BPH,traced under the headings of Waram unuq al-mathana(bladder neck swelling)and Insidad majra-i-mathana(bladder outlet obstruction),has been managed for centuries with herbal medicines yet demanding a comprehensive scientific validation.Among the herbs,Cucurbita pepo,Tribulus terrestris,Urtica dioica,and Linum usitatissimum are worth mentioning.Conclusion:For achieving the goal of LUTS-free ageing men,and safer and cost-effective future management of BPH,Unani herbal medicine could hopefully prove beneficial.